Field of the Invention
The present invention relates to the field of genetic engineering, and more particularly relates to a fused NHase with improved specific activity and stability.
Description of the Related Art
Nitrile hydratase (NHase; EC 4.2.1.84) is an enzyme catalyzing the hydration of a broad scope of nitriles to the corresponding amides. The NHase comprises a β-subunit, a α-subunit and a regulatory subunit and it is generally divided into the cobalt-type (Co-NHase) or the iron type (Fe-NHase) depending on the metal ion chelated with the active site.
NHase has been widely used in the industrial production of highly purified acrylamide and nicotinamide, since biotechnology synthesis has advantages of low-cost, low-energy consumption and less pollution compared to traditional chemical synthesis. However, most NHases with high activity are unstable during industrial application. For example, the NHases of Pseudomonas chlororaphils B23 and Rhodococcus sp. N-774 are unstable above 20° C., and the NHase of Rhodococcus rhodochrous J1 is merely stable between 10° C. and 30° C. In addition, it is necessary to maintain low reaction temperature to stabilize the NHases by refrigeration because of the exothermic reaction of nitrile-hydration, which usually causes enormous redundant energy cost. Furthermore, tolerance of NHase to high concentrations of the product is necessary in industrial manufacturing. Therefore, a more stable NHase with high activity and high tolerance is required for industrial manufacturing.